The effects of loading conditions and specimen environment on the nanomechanical response of canine cortical bone

Bone is a viscoelastic connective tissue composed primarily of mineral and type I collagen, which interacts with water, affecting its mechanical properties. Therefore, both the level of hydration and the loading rate are expected to influence the measured nanomechanical response of bone. In this study, we investigated the influence of three distinct hydration conditions, peak loads and loading/unloading rates on the elastic modulus and hardness of canine femoral cortical bone via nanoindentation. Sections from three canine femurs from multiple regions of the diaphysis were tested for a total of 670 indentations. All three hydration conditions (dry, moist and fully hydrated tissue) were tested at three different loading profiles (a triangular loading profile with peak loads of 600, 800 and 1000 μN at loading/unloading rate of 60, 80 and 100 μN/s, respectively; each test was 20s in duration). Significant differences were found for both the elastic modulus and hardness between the dry, moist and fully hydrated conditions (p≤0.02). For dry bone, elastic modulus and hardness values were not found to be significantly different between the different loading profiles (p>0.05). However, in both the moist and fully hydrated conditions, the elastic modulus and hardness were significantly different under all loading profiles (with the exception of the moist condition at the 600- and 800-μN peak load). Given these findings, it is critical to perform nanoindentation of bone under fully hydrated conditions to ensure physiologically relevant results. Furthermore, this work found that a 20-s triangular loading/unloading profile was sufficient to capture the viscoelastic behavior of bone in the 600- to 1000-μN peak load range. Lastly, specific peak load values and loading rates need to be selected based on the structural region for which the mechanical properties are to be measured.